The functions of thrombospondin in cell adhesion and migration are being investigated. We have identified two regions of the thrombospondin molecule that mediate adhesive and migratory responses of cultured human melanoma cells to thrombospondin. The carboxyl-terminal domain mediates attachment and haptotaxis, while the amino-terminal domain mediates cell spreading and chemotaxis. The cell receptors recognizing these two regions of thrombospondin are under investigation. one class of receptors are sulfated glycoconjugates which bind to the amino-terminal domain of thrombospondin. A minor heparin sulfate proteoglycan that binds thrombospondin with high affinity was identified in two melanoma cell lines and purified by affinity chromatography on thrombospondin-Sepharose (Cancer Res. 48: 6875, 1988). An unusual sulfated glycolipid present only in melanoma cell lines that spread on thrombospondin was also found to bind thrombospondin (ibid.). This glycolipid purified from peripheral nerve and a monoclonal antibody to the glycolipid specifically inhibit melanoma cell spreading on thrombospondin but not on fibronectin. To further define the mechanism of thrombospondin interactions with tumor cells, receptors for the carboxyl-terminus of thrombospondin will be characterized. Small cell lung carcinoma cells which attach on thrombospondin but not on other adhesive proteins will be used to identify specific thrombospondin receptors. The intracellular responses of cells to binding of thrombospondin to the two types of receptors are also being investigated.